The protozoan parasite Entamoeba histolytica causes an estimated 50 million cases of invasive disease annually and is the second leading parasitic cause of death worldwide. The most common manifestations of amebic infection are colonic disease (dysentery) and liver abscesses. In addition to the lives lost there is significant morbidity associated with amebic diarrhea! illness, especially in children who often suffer from malnourishment and growth retardation. E. histolytica is classified as a Class B agent of concern for bioterrorism and malicious use could result in significant morbidity. The parasite's life cycle involves conversion from the trophozoite to the cyst stage. This conversion, which is essential for disease propagation, is extremely poorly understood at present. Our goals are to develop tools to study E. histolytica developmental biology. The first aim is to develop a genome-wide transcriptional profile of E. histolytica cyst and trophozoites. The cyst transcriptional profiling will be performed using in vivo cysts and the associated excysted trophozoites and will provide the first genome- wide identification of cyst and trophozoite-specific genes. The second aim is to develop a method for in vitro encystation in E. histolytica. We will utilize strains of E. histolytica, which may be predisposed to encyst, grow them in conditions that mimic the colonic environment, and expose them to stimuli that cause encystation in related systems. Successful development of in vitro encystation will allow us to dissect key aspects of the developmental biology of this parasite including identification of the pathways that trigger encystation. These approaches promise to provide the first insights into differentiation in E. histolytica and will pave the way for further characterization of the development program in this parasite. Relevance to public health: Entamoeba histolytica is an important pathogen and has an impact on human health on a global scale. Although the majority of disease is in developing countries, this parasite can cause infections anywhere that water purification systems get adversely affected. Events such as the Tsunami or upheaval in the political and social infrastructure of a region can allow disease to emerge. We are interested in understanding the life stages of the parasite with the goal of identifying new ways to diagnose, treat, and prevent human infections. [unreadable] [unreadable] [unreadable]